Ecology and Chronic Wasting Disease Epidemiology Shape Prion Protein Gene Variation in Rocky Mountain Elk (Cervus elaphus nelsoni).

As chronic wasting disease (CWD) continues to spread across North America, the relationship between CWD and host genetics has become of interest. In Rocky Mountain elk (Cervus elaphus nelsoni), one or two copies of a leucine allele at codon 132 of the prion protein gene (132L*) has been shown to prolong the incubation period of CWD. Our study examined the relationship between CWD epidemiology and codon 132 evolution in elk from Wyoming, USA, from 2011 to 2018. Using PCR and Sanger sequencing, we genotyped 997 elk and assessed the relationship between genotype and CWD prevalence estimated from surveillance data. Using logistic regression, we showed that each 1% increase in CWD prevalence is associated with a 9.6% increase in the odds that an elk would have at least one copy of leucine at codon 132. In some regions, however, 132L* variants were found in the absence of CWD, indicating that evolutionary and epidemiologic patterns can be heterogeneous across space and time. We also provide evidence that naturally occurring CWD is not rare in 132L* elk, which merits the study of shedding kinetics in 132L* elk and the influence of genotype on CWD strain diversity. The management implications of cervid adaptations to CWD are difficult to predict. Studies that investigate the degree to which evolutionary outcomes are shaped by host spatial structure can provide useful epidemiologic insight, which can in turn aid management by informing scale and extent of mitigation actions.

Comparative Pathogenicity of Wildlife and Bovine Escherichia coli O157:H7 Strains in Experimentally Inoculated Neonatal Jersey Calves.

Shiga toxin-producing Escherichia coli, like E. coli O157:H7, are important human and animal pathogens. Naturally-acquired E. coli O157:H7 infections occur in numerous species but, particularly, cattle have been identified as a significant reservoir for human cases. E. coli O157:H7 are isolated from a number of domestic and wild animals, including rodents that share a living space with cattle. These Shiga toxin-producing E. coli O157:H7 strains can be highly virulent in humans, but little is known about the sequelae of interspecies transfer. In a group of neonatal calves, we determined the differences in colonization patterns and lesions associated with infection using either a wildlife or bovine E. coli O157:H7 strain. In calves challenged with the wildlife E. coli O157:H7 strain, the large (descending) colon was solely colonized, which differed substantially from the calves inoculated with the bovine E. coli O157:H7 strain, where the spiral colon was the principal target of infection. This study also demonstrated that while both interspecies- and intraspecies-derived E. coli O157:H7 can infect young calves, the distribution and severity differs.

Assessment of a fluorescent antibody test for the detection of antibodies against epizootic bovine abortion.

The current study was directed at developing and validating an indirect fluorescent antibody test (IFAT) capable of detecting antibodies specific for the agent of epizootic bovine abortion (aoEBA). Sensitivity and specificity was determined by comparing antibody titers from 114 fetuses infected with aoEBA with 68 fetuses diagnosed with alternate infectious etiologies. Data established specificity at 100% and sensitivity at 94.7% when cutoff criteria for a positive test were assigned at a titer of ≥1,000. Potential cross-reactivity was noted in samples from 3 fetuses with antibody titers of 10 or100; all were infected with Gram-positive organisms. The remaining 65 fetuses infected with microbes other than aoEBA, and an additional 12 negative reference sera, did not have detectable titers. The IFAT-based serology assay is rapid, reproducible, and unaffected by fluid color or opacity. Total fetal immunoglobulin (Ig)G was also evaluated as an aid for diagnosing EBA. Significantly higher concentrations of IgG were identified in fetuses infected with aoEBA as compared to those with alternate infectious etiologies. The presence of IgG is a sensitive indicator of EBA and increases the specificity of FAT-based serologic diagnosis when titers are 10 or 100. Taken together, serology and IgG analyses suggest that the incidence of EBA may be underestimated.

Evaluation of Anaplasma phagocytophilum infection in experimentally inoculated sheep and determination of Anaplasma spp seroprevalence in 8 free-ranging sheep flocks in California and Oregon.

OBJECTIVE: To evaluate disease progression in sheep experimentally inoculated with Anaplasma phagocytophilum and determine the Anaplasma spp seroprevalence in sheep in free-ranging flocks in the Sierra Nevada foothills and Oregon Coast Range. ANIMALS: 10 mature ewes seronegative for Anaplasma spp and 251 sheep from 8 flocks. PROCEDURES: 10 ewes received 1 of 3 treatments: A phagocytophilum Webster strain (n = 4), A phagocytophilum MRK strain (4), or human promyelocytic leukemia cells (control treatment [2]). Sheep were monitored for signs of clinical disease, and blood samples were obtained for serologic and PCR assay evaluation intermittently for 48 days. From a subsample of sheep from each of 8 free-ranging flocks, blood samples were obtained to determine Anaplasma spp seroprevalence. RESULTS: Sheep inoculated with A phagocytophilum developed subclinical or mild disease, whereas sheep inoculated with the control treatment did not develop any signs of disease. Only 2 ewes seroconverted; both had received the MRK strain. Anaplasma-specific DNA was detected in blood samples from 1 sheep in the Webster strain-inoculated group and 3 sheep in the MRK strain-inoculated group. Sheep seropositive for Anaplasma spp were detected in 5 of 8 flocks, and flocks in the Sierra Nevada foothills had higher within-flock seroprevalence (22%) than did flocks in the Oregon Coast Range (6.4%). CONCLUSIONS AND CLINICAL RELEVANCE: Infection with A phagocytophilum in mature sheep generally resulted in subclinical disease. Higher Anaplasma spp seroprevalence in sheep in the Sierra Nevada foothills corresponded to the geographic distribution of anaplasmosis reported for dogs, horses, and humans.

Effect of a nutritional reconditioning program for thin dairy cattle on body weight, carcass quality, and fecal pathogen shedding.

OBJECTIVE: To assess changes in body weight, carcass quality, and fecal pathogen shedding in cull dairy cows fed a high-energy ration for 28 or 56 days prior to slaughter. DESIGN: Randomized clinical trial. ANIMALS: 31 adult Holstein dairy cows. PROCEDURES: Cows were randomly assigned to a control (immediate slaughter) group or a 28-day or 56-day feeding group. Cows in the feeding groups received a high-energy feed and were weighed every 7 days. Carcasses were evaluated by USDA employees. Fecal and blood samples were collected at the start and end of the feeding periods. RESULTS: Body condition score and adjusted preliminary yield grade were significantly increased in both feeding groups, compared with values for the control group; body weight, hot carcass weight, dressing percentage, and ribeye area were significantly increased after 56 days, but not after 28 days, compared with values for the control group. Average daily gain and marbling score were significantly lower after feeding for 28 days versus after 56 days. Prevalence of Escherichia coli O157:H7 shedding in feces decreased from 14% to 5.6%, but this difference was not significant. Cows seropositive for antibodies against bovine leukemia virus that had signs of lymphoma and lame cows had a low average daily gain. Net loss was $71.32/cow and $112.80/cow for the 28-day and 56-day feeding groups, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: Feeding market dairy cows improved body condition and carcass quality. Cows seropositive for antibodies against bovine leukemia virus that have signs of lymphoma and lame cows might be poor candidates for reconditioning.

Prevalence and molecular characterization of Escherichia coli O157:H7 by multiple-locus variable-number tandem repeat analysis and pulsed-field gel electrophoresis in three sheep farming operations in California.

A year-long study was conducted to determine the fecal prevalence of Escherichia coli O157:H7 in three sheep ranches. Strain diversity and persistence were compared with multiple-locus variable-number tandem repeat analysis and pulsed-field gel electrophoresis. Ranch C, a feedlot, consisted of young sheep raised predominantly on a high-grain diet. The other two sites consisted of sheep raised on native pasture and a combination of native and irrigated pasture. Forty fecal samples were collected every month from each ranch. Samples were examined for E. coli O157:H7 by immunomagnetic separation and culture of the magnetic beads onto selective media. Detection of virulence markers in positive isolates was determined by PCR. E. coli O157:H7 was isolated from 100 (22.7%) of 440 fecal samples collected from ranch C. On ranch B, 9 (1.9%) of the 480 fecal samples were positive for the pathogen, while none of the samples from ranch A were positive. On ranch C, the odds of detecting E. coli O157:H7 was 3.2 times greater during the warmer months compared with the cooler months of the year. There was no association between days spent in the feedlot and fecal prevalence of the pathogen (P = 0.62). Most multiple-locus variable-number tandem repeat analysis types were isolated only once from ranch C (14 of 23), but several strains were isolated over 4 to 6 months, often in many intervening negative months. This study revealed that the prevalence of E. coli O157:H7 can be high in some sheep ranches in California, especially in feedlots where young sheep are fed predominantly high-grain rations.

Longitudinal prevalence and molecular typing of Escherichia coli O157:H7 by use of multiple-locus variable-number tandem-repeat analysis and pulsed-field gel electrophoresis in fecal samples collected from a range-based herd of beef cattle in California.

OBJECTIVE: To evaluate seasonal patterns and risk factors for Escherichia coli O157:H7 in feces in a beef cattle herd and determine strain diversity and transition in E coli over time by use of multiple-locus variable-number tandem-repeat analysis (MLVA) and pulsed-field gel electrophoresis (PFGE). SAMPLE POPULATION: 456 samples of freshly passed feces collected over a 1-year period from cattle in a range-based cow-calf operation located in the foothills of the Sierra Nevada Mountains in California. PROCEDURES: E coli O157:H7 was recovered from feces by use of immunomagnetic separation and 2 selective media. Virulence factors were detected via reverse transcriptase-PCR assay. Escherichia coli O157:H7 isolates were subtyped with MLVA and PFGE. Prevalence estimates were calculated and significant risk factors determined. A dendrogram was constructed on the basis of results of MLVA typing. RESULTS: Overall prevalence estimate for E coli O157:H7 was 10.5%, with the prevalence lowest during the winter. Mean temperature during the 30 days before collection of samples was significantly associated with prevalence of E coli O157:H7 in feces. Nineteen MLVA and 12 PFGE types were identified. CONCLUSIONS AND CLINICAL RELEVANCE: A seasonal pattern was detected for prevalence of E coli O157:H7 in feces collected from beef cattle in California. Subtyping via MLVA and PFGE revealed a diversity of E coli O157:H7 strains in a cow-calf operation and noteworthy turnover of predominant types. Given the importance of accurately determining sources of contamination in investigations of disease outbreaks in humans, MLVA combined with PFGE should be powerful tools for epidemiologists.

Pathology associated with West Nile virus infections in the yellow-billed magpie (Pica nuttalli): a California endemic bird.

Yellow-billed Magpies (Pica nuttalli, Corvidae) are found exclusively in central California and have experienced alarming West Nile virus (WNV)-associated mortality since 2004. The first reported case of WNV in the species was reported in July 2004. Subsequently, 81% (304/374) of dead magpies submitted that year to the California Department of Health Services Dead Bird Surveillance Program were WNV positive by polymerase chain reaction (PCR) test. We studied 43 magpie carcasses collected in 2004 and observed distinctive lesions in 24 birds that tested positive for WNV. Lesions included vasculitis and necrosis, and organs affected included brain, heart, liver, and gastrointestinal tract. From the severity of lesions observed, we suspect that a rapid onset of morbidity and mortality occurs with the Yellow-billed Magpie. Examination of bird survey data indicates that Yellow-billed Magpie abundance declined coincidentally with the onset of WNV in California. The home range and habitat of the species are nested within known areas of WNV transmission. Yellow-billed Magpies may be at risk of a decline and population bottleneck. Observations and experience with the Yellow-billed Magpie and WNV may provide insights for other endangered corvids that have not yet been exposed to WNV, including the Hawaiian Crow (Corvus hawaiiensis) and Island Scrub Jay (Aphelocoma insularis).

Molecular genealogy tools for white-tailed deer with chronic wasting disease.

Molecular genetic data provide powerful tools for genealogy reconstruction to reveal mechanisms underlying disease ecology. White-tailed deer (Odocoileus virginianus) congregate in matriarchal groups; kin-related close social spacing may be a factor in the spread of infectious diseases. Spread of chronic wasting disease (CWD), a prion disorder of deer and their cervid relatives, is presumed to be associated with direct contact between individuals and by exposure to shared food and water sources contaminated with prions shed by infected deer. Key aspects of disease ecology are yet unknown. DNA tools for pedigree reconstruction were developed to fill knowledge gaps in disease dynamics in prion-infected wild animals. Kinship indices using data from microsatellite loci and sequence haplotypes of mitochondrial DNA were employed to assemble genealogies. Molecular genealogy tools will be useful for landscape-level population genetic research and monitoring, in addition to epidemiologic studies examining transmission of CWD in captive and free-ranging cervids.

Giardia duodenalis in feedlot cattle from the central and western United States.

BACKGROUND: Giardia duodenalis is a ubiquitous protozoan parasite that has emerged as a significant opportunistic human pathogen. G. duodenalis may have a deleterious effect on animal growth and performance, therefore its potential as a production limiting organism should not be discounted. We therefore undertook this study to determine management and environmental factors in feedlots that influence the prevalence and environmental load of G. duodenalis cysts in fecal material deposited by feedlot cattle in the central and western United States. RESULTS: Twenty two feedlots from 7 states were included in the study, and up to 240 fecal samples were collected from pen floors of up to 6 pens per feedlot. Giardia duodenalis cysts were identified and counted using direct immunofluorescent microscopy. The estimated overall point prevalence of G. duodenalis was 19.1%, representing feedlots from a wide range of climates and management systems. Pen-level prevalence varied from 0 to 63.3%, with pen-level shedding estimates ranging from 0 to 261,000 cysts/g feces. Higher environmental temperatures, increased animal density, and increased time in the feedlot were associated with a lower prevalence of G. duodenalis. Removing manure before placing a new group of cattle in a pen was associated with a decreased prevalence of G. duodenalis in fecal pats. Using coccidiostats as a feed additive was associated with a higher prevalence of Giardia. CONCLUSION: Management practices could be employed that would limit the probability that feedlot cattle shed G. duodenalis in their feces and therefore potentially limit contamination of their environment.

Evaluation of sequential coinfection with Anaplasma phagocytophilum and Anaplasma marginale in cattle.

OBJECTIVE: To determine whether sequelae of infection differed among single versus double infection with Anaplasma phagocytophilum or Anaplasma marginale, with and without tick salivary extract, in cattle. ANIMALS: Eighteen 13-month old steers. PROCEDURES: Treatment groups of 3 cattle each included A marginale inoculated ID followed on day 35 by A phagocytophilum without tick saliva, A phagocytophilum followed on day 10 by A marginale without tick saliva, A marginale followed on day 35 by A phagocytophilum with tick saliva, A phagocytophilum followed on day 10 by A marginale with tick saliva, tissue culture control injection, and tick saliva control injection. Infection was monitored via clinical observations, CBC, serologic testing, and PCR analysis of blood and tissues. RESULTS: Infected cattle had significantly reduced weight gain. Anemia occurred 25 to 32 days after A marginale infection, which was attenuated by tick saliva. Parasitism was greater if cattle had not previously been inoculated with A phagocytophilum. Nine of the 12 treated cattle had positive results of PCR analysis for A phagocytophilum from at least 1 blood sample. Five tissue samples had positive results of PCR analysis for A phagocytophilum; PCR results for A marginale were positive in spleen, lung, lymph node, heart, and ear skin of infected cattle. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated an important biological interaction between A marginale and A phagocytophilum infection as well as with tick saliva in disease kinetics and severity in cattle, which may be important for interpretation of diagnostic tests and management of disease in areas where both pathogens occur.

Evaluation of twice-daily, low-dose trilostane treatment administered orally in dogs with naturally occurring hyperadrenocorticism.

OBJECTIVE: To evaluate the effects of twice-daily oral administration of a low-dose of trilostane treatment and assess the duration of effects after once-daily trilostane administration in dogs with naturally occurring hyperadrenocorticism (NOH). DESIGN: Prospective study. ANIMALS: 28 dogs with NOH. PROCEDURES: 22 dogs received 0.5 to 2.5 mg of trilostane/kg (0.23 to 1.14 mg/lb) orally every 12 hours initially. At intervals, dogs were reevaluated; owner assessment of treatment response was recorded. To assess drug effect duration, 16 of the 22 dogs and 6 additional dogs underwent 2 ACTH stimulation tests 3 to 4 hours and 8 to 9 hours after once-daily trilostane administration. RESULTS: After 1 to 2 weeks, mean trilostane dosage was 1.4 mg/kg (0.64 mg/lb) every 12 hours (n = 22 dogs; good response [resolution of signs], 8; poor response, 14). Four to 8 weeks later, mean dosage was 1.8 mg/kg (0.82 mg/lb) every 12 or 8 hours (n = 21 and 1 dogs, respectively; good response, 15; poor response, 5; 2 dogs were ill). Eight to 16 weeks after the second reevaluation, remaining dogs had good responses (mean dosages, 1.9 mg/kg [0.86 mg/lb], q 12 h [n = 13 dogs] and 1.3 mg/kg [0.59 mg/lb], q 8 h [3]). At 3 to 4 hours and 8 to 9 hours after once-daily dosing, mean post-ACTH stimulation serum cortisol concentrations were 2.60 and 8.09 Pg/dL, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: In dogs with NOH, administration of trilostane at low doses every 12 hours was effective, although 2 dogs became ill during treatment. Drug effects diminished within 8 to 9 hours. Because of potential adverse effects, lower doses should be evaluated.

The correlation of potassium content and moisture in bovine liver samples analyzed for trace mineral concentrations.

Beef cattle in the United States are often found to be deficient in essential trace minerals such as copper and zinc. Established reference ranges for mineral concentrations exist and usually designate a concentration as adequate, marginal, deficient, or excessive. This research investigates a new method of interpreting detected elemental concentrations in bovine liver that will add confidence to the final diagnosis. This is based on the hypothesis that a correlation exists between potassium concentration and moisture in a bovine liver sample. This relationship between potassium and moisture content enables the diagnostician to more accurately predict mineral concentrations and wet weight regardless of sample moisture loss. Correlations were found between potassium content and percentage of moisture in experimental samples, clinical biopsies, and a validation study, to a statistical significance of P < 0.001. Experimental samples had a correlation coefficient of R2 = 0.95 and the mathematical relationship y = 2513.2x(-1.0662). Clinical biopsies had a correlation of R2 = 0.83 and the mathematical relationship y = 2203.4x(-0.991). The validation study had a correlation of R2 = 0.55 and a mathematical relationship y = 2321.4x(-0.952). An exponent of -1 is predicted by conservation of potassium mass. These findings have practical significance in maintaining and improving cattle growth, health, reproduction, and food safety.

Vitamin E and selenium concentrations in month-old beef calves.

Three groups of beef cow and calf pairs were studied to determine plasma vitamin E and blood selenium (Se) concentrations of calves at 1 month old. Group 1 was managed on irrigated pasture and calves received no Se/vitamin E injections at birth. Group 2 was managed on irrigated pasture, and the calves were injected with Se/vitamin E at birth. Group 3 was managed on dry foothill grasslands, and these cows were supplemented with 56.3 mg vitamin E and 3 mg Se daily, and the calves received a Se/vitamin E injection at birth. The plasma concentration of vitamin E in group 1 and 2 cows (9.5 +/- 1.24 and 8.43 +/- 1.0 microg/ml, respectively) was significantly higher than that of the group 3 cows (2.28 +/- 0.42 microg/ml; P < 0.05). The blood Se concentrations in group 3 cows (169 +/- 37 ng/ml) were significantly higher than those in group 1 and 2 cows (36.4 +/- 15.9 and 31.1 +/- 12.5 ng/ml, respectively; P < 0.05). Calf Se was highly correlated to cow Se (r = 0.965), and calf vitamin E was moderately correlated to cow vitamin E (r = 0.605). Calf vitamin E concentrations were consistently lower than cow vitamin E concentrations, and many values would be considered deficient.

Prevalence of Neospora caninum and persistent infection with bovine viral diarrhea virus in dairy-breed steers in a feedlot.

OBJECTIVE: To determine the prevalence and effect of Neospora caninum infection and persistent infection (PI) with bovine viral diarrhea virus (BVDV) on weight gain, morbidity, and mortality rate in dairy-breed steer calves located on a feedlot in California. DESIGN: Prospective cohort observational study. ANIMALS: 900 dairy-breed steer calves in 2 pens. PROCEDURES: The 3- to 4-month-old calves were evaluated for serum antibodies against N caninum and infection with BVDV at entry to the feedlot. Five months later, sera were again analyzed for anti-N caninum antibodies; calves that were determined to have BVDV infection initially were retested to evaluate PI status. Average daily gain, morbidity, and deaths were recorded for all calves. RESULTS: Among 900 calves, prevalence of N caninum infection was 16.7% (95% confidence interval, 14.3% to 19.3%); prevalence of BVDV-associated PI was 0.2% (95% confidence interval, 0.03% to 0.9%). Morbidity rate and time to first illness were not significantly different between calves that were seropositive or seronegative for N caninum. At the second sample collection, weight and average daily gain of calves that were seropositive for N caninum was less than that of seronegative steers in 1 pen, whereas these measures did not differ between groups in the other pen. Statistical power was insufficient to evaluate the effect of BVDV PI on any outcome measurement. CONCLUSIONS AND CLINICAL RELEVANCE: Although N caninum serostatus had no significant effect on morbidity rate, some seropositive calves had reduced growth, compared with seronegative calves, 5 months after entry to the feedlot.

Probability of introduction of exotic strains of bluetongue virus into the US and into California through importation of infected cattle.

Strategies designed to minimize the probability of bluetongue virus (BTV) introduction to new areas should be based on a quantitative assessment of the probability of actually establishing the virus once it is introduced. The risk of introducing a new strain of bluetongue virus into a region depends on the number of viremic animals that enter and the competency of local vectors to transmit the virus. We used Monte Carlo simulation to model the probability of introducing BTV into California, USA, and the US through importation of cattle. Records of cattle and calf imports into California and the US were obtained, as was seroprevalence information from the exporting countries. A simulation model was constructed to evaluate the probability of importing either a viremic PCR-negative animal after 14-day quarantine, a c-ELISA BTV-antibody-negative animal after 28-day quarantine, or an untested viremic animal after 100-day quarantine into California and into the US. We found that for animals imported to the US, the simulated (best to worst scenarios) median percentage that tested positive for BTV-antibody ranged from 5.4 to 7.2%, while for the subset imported to California, the simulated median percentage that tested positive for BTV-antibody ranged from 20.9 to 78.9%. Using PCR, for animals imported to the US these values were 71.8-85.3%, and for those imported to California, the simulated median that test positive ranged from 74.3 to 92.4%. The probability that an imported animal was BTV-viremic is very low regardless of the scenario selected (median probability=0.0%). The probability of introducing an exotic strain of BTV into California or the US by importing infected cattle was remote, and the current Office International des Epizooties (OIE) recommendation of either a final PCR test performed 14 days after entry into quarantine, a c-ELISA performed 28 days after entry into quarantine or a 100-day quarantine with no testing requirement was adequate to protect cattle in the US and California from an exotic strain of BTV.

Regional risk of exporting cattle seropositive for bluetongue virus from the United States.

OBJECTIVE: To create a stochastic model to quantify the risk that shipments of cattle from regions within the United States would contain animals seropositive for bluetongue virus and to determine shipment-level accuracy of serologic testing by use of a competitive ELISA (c-ELISA). SAMPLE POPULATION: 19,216 shipments containing 528,918 cattle and calves. PROCEDURE: Data were obtained on number of animals and state of origin of cattle in export shipments originating within the United States between January 1994 and March 2002. Probability distributions for size of export shipments were determined for all states within the United States, and distributions for agar gel immunodiffusion and c-ELISA accuracy (sensitivity and specificity) were determined from expert opinion and review of the literature. The model simulated selection of a shipment and then determined the probability that a threshold number or percentage of cattle within that shipment would have a positive c-ELISA result. Shipment-level sensitivity, specificity, positive-predictive value, and negative-predictive value were calculated. RESULTS: Substantial differences were evident in the regional probability of a shipment being declared positive, with shipments from northeastern states having the lowest probability and shipments from southwestern states having the highest probability. The c-ELISA had variable predictive values at the shipment level, depending on the threshold used and the prevalence of antibody-positive cattle within the region. CONCLUSIONS AND CLINICAL RELEVANCE: Results from this study will aid importers in making scientifically based decisions regarding risk of importing antibody-positive cattle.